NMR Derivatives for Quantification of<sup>2</sup>H and<sup>13</sup>C‐Enrichment of Human Glucuronide from Metabolic Tracers

Jones, John G.; Barosa, Cristina; Gomes, Filipe; Mendes, Ana Carina Loureiro; Delgado, Teresa C.; Diogo, Luísa; Garcia, Paula; Bastos, Maria de Lourdes; Barros, Luísa; Fagulha, Ana; Baptista, Carla; Carvalheiro, Manuela; Caldeira, M.Madalena

doi:10.1080/07328300600732840

Cited by 36 publications

(33 citation statements)

References 22 publications

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“…Sample processing and NMR spectroscopy. Acetaminophen glucuronide was converted to MAG as described (17). Proton-decoupled MAG 2 H NMR spectra were acquired at 11.75 T with a Varian Unity 500 system equipped with a 5-mm broadband probe (Varian, Palo Alto, CA).…”

Section: Methodsmentioning

confidence: 99%

Noninvasive Analysis of Hepatic Glycogen Kinetics Before and After Breakfast with Deuterated Water and Acetaminophen

et al. 2006

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The contributions of hepatic glycogenolysis to fasting glucose production and direct pathway to hepatic glycogen synthesis were quantified in eight type 1 diabetic patients and nine healthy control subjects by ingestion of 2 H 2 O and acetaminophen before breakfast followed by analysis of urinary water and acetaminophen glucuronide. After overnight fasting, enrichment of glucuronide position 5 relative to body water (G5/body water) was significantly higher in type 1 diabetic patients compared with control subjects, indicating a reduced contribution of glycogenolysis to glucose production (38 ؎ 3 vs. 46 ؎ 2%). Following breakfast, G5/body water was significantly higher in type 1 diabetic patients, indicating a smaller direct pathway contribution to glycogen synthesis (47 ؎ 2 vs. 59 ؎ 2%). Glucuronide hydrogen 2 enrichment (G2) was equivalent to body water during fasting (G2/body water 0.94 ؎ 0.03 and 1.02 ؎ 0.06 for control and type 1 diabetic subjects, respectively) but was significantly lower after breakfast (G2/body water 0.78 ؎ 0.03 and 0.82 ؎ 0.05 for control and type 1 diabetic subjects, respectively). The reduced postprandial G2 levels reflect incomplete glucose-6-phosphate-fructose-6-phosphate exchange or glycogen synthesis from dietary galactose. Unlike current measurements of human hepatic glycogen metabolism, the 2 H 2 O/acetaminophen assay does not require specialized on-site clinical equipment or personnel. Diabetes

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Section: Methodsmentioning

confidence: 99%

Noninvasive Analysis of Hepatic Glycogen Kinetics Before and After Breakfast with Deuterated Water and Acetaminophen

et al. 2006

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“…Paracetamol glucuronide in this solution was derivatized to monoacetoneglucuronic lactone (MAGL) as previously described (14). Two thousand units of ␤-glucuronidase (H. Pomatia, Sigma Chemical, St. Louis, MO) were added and the solution was incubated at 45°C for 48 hr.…”

Section: Sample Processingmentioning

confidence: 99%

Quantification of hepatic transaldolase exchange activity and its effects on tracer measurements of indirect pathway flux in humans

Jones¹,

Garcia²,

Barosa³

et al. 2008

Magnetic Resonance in Med

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Exchange of hepatic glucose-6-phosphate (G6P) and glyceraldehyde-3-phosphate via transaldolase modifies hepatic G6P enrichment from glucose or gluconeogenic tracers. Transaldolase exchange was quantified in five healthy, fed subjects following an oral bolus of [1,2,3-13 C 3 ]glycerol (25-30 mg/kg) and paracetamol (10 -12 mg/kg). 13 Hepatic glucose-6-phosphate (G6P) lies at the metabolic crossroads of hepatic glucose and glycogen metabolism. Under fasting conditions when the liver is a net producer of glucose, hepatic G6P is generated by the hydrolysis of glycogen and from gluconeogenesis and is then converted to glucose via G6P. Under fed conditions there is net hepatic glycogen synthesis from both glucose and from gluconeogenic precursors. These metabolic pathways converge at G6P and the hexose carbon skeletons are then incorporated into glycogen via glucose-1-phosphate and UDP-glucose. The gluconeogenic contribution to G6P synthesis is modified in a variety of human diseases, including insulin-and noninsulin-dependent diabetes, cirrhosis, and malaria (1-4). Therefore, quantifying the fraction of G6P derived from gluconeogenesis is a key parameter for defining hepatic carbohydrate metabolism under these and other pathophysiological conditions. In humans, several different tracer methods have been developed for quantifying the contribution of gluconeogenesis to hepatic G6P flux. Hepatic G6P enrichment from these tracers can be quantified noninvasively by analysis of urinary glucuronide enrichment (5-7).Implicit in all measurements of hepatic gluconeogenesis using labeled gluconeogenic substrates is the assumption that G6P molecules derived from non-gluconeogenic precursors (i.e., glycogenolysis) are not labeled with the tracer. 1 Likewise, when labeled glucose is used to determine the contribution of direct and indirect pathways of hepatic glycogen synthesis under fed conditions, dilution of the glucose tracer at the level of hepatic G6P is assumed to be entirely due to gluconeogenic G6P production.The possibility that transaldolase (TA) exchange could invalidate these assumptions was recognized by Landau and co-workers (8 -10). TA catalyzes the exchange between the glyceraldehyde-3-phosphate (GA3P) moiety (i.e., carbons 4, 5, and 6) of fructose-6-phosphate (F6P) and free GA3P in many tissues (11)(12)(13). This exchange is independent of oxidative pentose phosphate pathway (PPP) flux, hence tissues that have relatively low oxidative PPP utilization of G6P, such as liver, may nevertheless have significant TA exchange activity (13). Since G6P and F6P are in rapid exchange, G6P molecules derived from glucose or glycogen are exposed to TA activity. With gluconeogenic tracers that label GA3P, the effect of TA exchange is to transfer the label to G6P molecules derived from glucose or glycogen. Hence, the gluconeogenic contribution is overestimated relative to the contribution from

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“…The acetic acid fractions were evaporated off, and the residue was dissolved in 0.1 mol/l acetate buffer, pH 5.0. The acetaminophen glucuronide was converted to monoacetoneglucose (MAG) by a slight modification of the method of Jones et al (21). Acetaminophen glucuronide in acetate buffer was added with glucuronidase (Helix Pomatia) and incubated for 48 h at 37°C.…”

Section: Diabetes Vol 57 January 2008mentioning

confidence: 99%

Evidence That Processes Other Than Gluconeogenesis May Influence the Ratio of Deuterium on the Fifth and Third Carbons of Glucose

et al. 2008

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OBJECTIVE-The deuterated water method uses the ratio of deuterium on carbons 5 and 2 (C5/C2) or 3 and 2 (C3/C2) to estimate the fraction of glucose derived from gluconeogenesis. The current studies determined whether C3 and C5 glucose enrichment is influenced by processes other than gluconeogenesis.RESEARCH DESIGN AND METHODS-Six nondiabetic subjects were infused with [3,5-2 H 2 ]glucose and insulin while glucose was clamped at ϳ5 mmol/l; the C5-to-C3 ratio was measured in the in UDP-glucose pool using nuclear magnetic resonance and the acetaminophen glucuronide method.RESULTS-Whereas the C5-to-C3 ratio of the infusate was 1.07, the ratio in UDP-glucose was Ͻ1.0 in all subjects both before (0.75 Ϯ 0.07) and during (0.67 Ϯ 0.05) the insulin infusion.CONCLUSIONS-These data indicate that the deuterium on C5 of glucose is lost more rapidly relative to the deuterium on C3. The decrease in the C5-to-C3 ratio could result from exchange of the lower three carbons of fructose-6-phosphate with unlabeled three-carbon precursors via the transaldolase reaction and/or selective retention of the C3 deuterium at the level of triosephosphate isomerase due to a kinetic isotope effect. After ingestion of 2 H 2 O, these processes would increase the enrichment of C5 and decrease the enrichment of C3, respectively, with the former causing an overestimation of gluconeogenesis using the C2-to-C5 ratio and the latter an underestimation using the C3-to-C2 ratio. Future studies will be required to determine whether the impact of these processes on the measurement of gluconeogenesis differs among the disease states being evaluated (e.g., diabetes or obesity). Diabetes 57:50-55, 2008

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NMR Derivatives for Quantification of²H and¹³C‐Enrichment of Human Glucuronide from Metabolic Tracers

Cited by 36 publications

References 22 publications

Noninvasive Analysis of Hepatic Glycogen Kinetics Before and After Breakfast with Deuterated Water and Acetaminophen

Noninvasive Analysis of Hepatic Glycogen Kinetics Before and After Breakfast with Deuterated Water and Acetaminophen

Quantification of hepatic transaldolase exchange activity and its effects on tracer measurements of indirect pathway flux in humans

Evidence That Processes Other Than Gluconeogenesis May Influence the Ratio of Deuterium on the Fifth and Third Carbons of Glucose

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NMR Derivatives for Quantification of2H and13C‐Enrichment of Human Glucuronide from Metabolic Tracers

Cited by 36 publications

References 22 publications

Noninvasive Analysis of Hepatic Glycogen Kinetics Before and After Breakfast with Deuterated Water and Acetaminophen

Noninvasive Analysis of Hepatic Glycogen Kinetics Before and After Breakfast with Deuterated Water and Acetaminophen

Quantification of hepatic transaldolase exchange activity and its effects on tracer measurements of indirect pathway flux in humans

Evidence That Processes Other Than Gluconeogenesis May Influence the Ratio of Deuterium on the Fifth and Third Carbons of Glucose

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NMR Derivatives for Quantification of²H and¹³C‐Enrichment of Human Glucuronide from Metabolic Tracers